ABSTRACT The long-term goal of the proposed research is to understand the mechanisms that regulate organogenesis. The overall objective of the proposal is to use our novel in vivo force transducers, which allow quantitative measurements of mechanical stresses within living embryonic tissues, to unveil the role of mechanical signals in the specification of signaling centers and cell types during organ morphogenesis, using the tooth as a model system. The central hypothesis is that the endogenous regional variations in compressive and tensile stresses in the tissue control the distribution of nuclear YAP localization in the developing tooth, thereby regulating specification of key signaling centers. The following Specific Aims will employ a combination of novel technologies designed to measure mechanical stresses in vivo and in situ with sophisticated mouse genetic strategies. Aim 1 will characterize regional differences in endogenous compressive and tensile stresses during tooth development. These experiments will constitute the first ever measurement of regional differences in compressive and tensile stresses during the formation of any vertebrate organ. These regional changes in mechanics will be related to spatial variations in YAP nuclear localization in the tissue and the establishment of signaling centers. Aim 2 will determine the molecular control of signaling center formation by tensile and compressive stresses in the developing tooth in vivo. In order to link the in vivo stress measurements to the molecules controlling the mechanical phenotype, we will first image the spatiotemporal distribution of proteins involved in force generation. Moreover, we will genetically delete the genes encoding these proteins and determine how mutations in these genes affect YAP localization and the ability to generate compressive and tensile stresses in the tissue. Aim 3 will reveal the role of mechanical stresses in the regulation of nuclear vs. cytoplasmic YAP localization in vivo. These experiments will directly test our hypothesis that regional differences in compressive and tensile stresses in the tissue control the tissue distribution of nuclear YAP localization. Together, these studies will provide a leap forward in our knowledge of how tooth development is regulated by a novel signal, mechanical stress. Such information about the fundamental biology of tooth development will in turn enhance future efforts in applications such as tooth bioengineering.